The instant invention relates to the detection of Human Immunodeficiency Virus (HIV) in a biological sample containing bodily fluids, such as serum and plasma, tissues, or cell culture fluid and the like, utilizing an immunological capture assay involving the HIV nucleocapsid protein, p7, or an immunoreactive fragment of p7. Uses for this invention include determining the prognosis of disease in an HIV-infected person, monitoring the effectiveness of antiviral treatment, detecting HIV-infection in infants born to HIV-infected mothers, and detecting and quantitating HIV in laboratory experiments (i.e., virus production, infectivity assays, neutralization assays, drug effectiveness assays, etc.).
Surrogate markers of AIDS progression are needed to establish the requirement and effectiveness of any particular antiviral treatment. Several surrogate markers for disease progression have been studied, including measurement of absolute numbers of CD4+cells, .beta.-2 microglobulin plasma concentrations, serum neoptrin levels, amounts of infectious virus in PBLs, HIV-1 antigen levels in plasma and decrease or disappearance of antibodies to p24 [Jacobsen et al, British Medical Journal, 302:73 (1991) and Bagasra et al., N. Eng. J. Med., 326:1385-91 (1992)]. Difficulties exist with most of these markers when used to predict disease progression or to detect HIV infection, such as in infants born to HIV-infected mothers. Although there is a direct correlation between absolute numbers of CD4 positive T-cells and opportunistic infections resulting in death, CD4 cell counts are not satisfactory as disease progression markers because the slope of their decrease is low in most seropositive individuals [Andrieu, Clin. Exp. Immunol., 73:1 (1988)].
However, a direct correlation has been reported between the levels of infectious HIV-1 in plasma from infected individuals and AIDS progression, emphasizing the need to quantitatively determine HIV-1 in plasma of infected persons [Michael et al., J. Vir., 66:310-316 (1992); Katzenstein et al., J. Acquired. Imm. Deficiency Syndromes, 5:107-112 (1992) and Henrard et al., AIDS Res. & Human Retroviruses, 8:47 (1992)]. Increased concentrations of HIV-1 in plasma or serum is usually measured by (1) tissue culture assays to detect infectious virus (2) polymerase chain reaction (PCR) to detect the viral genomic RNA (3) antigen capture assays using the capsid antigen, p24, or matrix antigen, p17, to detect HIV-1 proteins. However, viral isolation and polymerase chain reaction assays are laborious, time consuming and require a very high level of technical competence. Detection of HIV-1 in plasma by p24 and p17 capture assays is not reliable because antibodies to these proteins in the patient interfere with detection in the capture assays [Vjhelyi et al., AIDS, 1:161-165 (1987) and Nishanian et al, J. Inf. Diseases, 962:21-28 (1990)]. Consequently p24 cannot be detected in many of the asymptomatic individuals and AIDS patients. Additionally, because of interference with maternal antibodies, p24 antigen capture assays are insensitive to detecting HIV infection in infants born of HIV-infected mothers [Cassol, et al., J. Acquired Imm. Deficiency Syndromes, 5:113-119 (1992)].
The importance of antigen capture assays to identify viremia is underscored by the development of various procedures employed to detect antigen in the presence of antibodies. These procedures include methods to disrupt immune complexes between anti-p24 and p24, [Mathiesen et al, J. Vir. Methods, 22:143-148 (1988); Fiscus et al., J. Infi Diseases, 164:765-9 (1991) and Kestens et al., J. Vir. Methods, 31:67-76 (1991)], measurement of the quantitative capacity for serum to complex with p24 [Crosxon et al, AIDS Research & Human Retroviruses, 6:455 (1990)] and use of antibodies to HIV-1 envelope antigens to capture the virus with subsequent detection by PCR [Henrard et al., AIDS Research & Human Retroviruses, 8:47 (1992)]. These more complicated procedures are limited to use in well-equipped laboratories and may suffer from sensitivity and reproducibility problems.
However, a simple, rapid assay to quantitate HIV-1 viremia would be valuable, for instance, as (1) a prognostic indicator to determine when an HIV-infected person is progressing to AIDS, to determine if an individual is infected with HIV, and to compliment the current antibody-based assays to detect HIV infection, (2) in monitoring the effectiveness of anti-viral treatment (or effectiveness of a new potential anti-HIV drug), (3) to determine if a vaccinated individual (i.e., a person that has been immunized with HIV, developed antibodies to HIV proteins and would consequently score positive in the antibody-based assays to detect HIV infection) was infected with HIV, (4) in determining if infants born to HIV-infected mothers are infected with HIV to allow early intervention and (5) as a method to detect and quantitate HIV in assays in clinical and research laboratories (e.g., isolation of HIV from PBMCs, propagation of HIV in cells, neutralization assays, drug-sensitivity assays, etc.) For instance, there is currently no simple test to predict or to confirm the presence of virus in an infant born to an HIV-1 infected mother. Neonatal IgM antibodies are often absent, p24 capture assays are insensitive and isolation of virus is slow, tedious, and requires specialized tissue culture facilities. However, detection is needed to permit early and effective clinical management of infected children and to reliably identify uninfected children. Thus, there exists a need for a surrogate marker for detecting HIV viremia and for a rapid, accurate assay for measurement of the levels of infectious HIV in the plasma, serum or other body fluids such as saliva and or cells such as PBMC's of infected individuals.
Furthermore, similar problems exist for lentiviruses other than HIV-1. Thus, there also exists a need for an accurate method for detecting and quantitating levels of HIV-2, SIV and other related lentivruses in biological samples obtained from animals.
Throughout this application, various publications are referenced. The disclosures of these publications in their entireties are hereby incorporated by reference into this application in order to more fully describe the state of the art to which this invention pertains.